Abstract The goal of this project is to develop IV injectable fluorescent agents for use in intraoperative imaging of peritoneal metastatic tumors. Significance: Peritoneal metastatic cancers have an incidence of 250,000/per year in the US and are primarily treated with surgery. Complete surgical tumor resection is a gold-standard treatment goal that can increase the 5-year patient survival to 50%, whereas incomplete tumor resection (<2mm) could drastically reduce the survival to a mere few months. Current clinical imaging technologies such as PET, MRI and CT have limitations for guiding cancer surgery including inability to be used intraoperatively due to safety associated with ionizing radiation, as well as limitations on sensitivity and tumor specificity especially for lesions smaller than 10mm. Conventional fluorescent molecular probes in development typically target tumor proteins and are limited in imaging the heterogeneous onco-geno-phenotypes of peritoneal metastasis that have multiple tissues of origin and vary in protein expression levels. In order to reduce incomplete tumor resection and improve survival, there is an urgent need for clear visual contrast imaging agents that enable more precise and specific detection of peritoneal metastasis tumors intraoperatively and in real-time. Hypothesis: OncoNano?s approach of targeting tumor acidosis which is ubiquitously expressed as a cancer biomarker across tumors, coupled with OncoNano?s ultra-sensitive pH fluorescent digital sensors, provides a broad based approach to image the heterogeneous peritoneal metastasis with high specificity and sensitivity. Preliminary Data: We have demonstrated that OncoNano?s fluorescent agents image tumors across a variety of solid tumor types (14 tested to date), including peritoneal metastasis to submillimeter precision in SCID mouse models. OncoNano?s agents provided a digital (ON-OFF) response which offered clear tumor delineation, and lower detection limits (>10x) compared with state of the art PET-FDG imaging. Surgical tumor resection using OncoNano imaging agent showed significant (40-50%) survival benefit in SCID mouse models compared with conventional white light based surgery. Specific Aims: The scientific aims of this project will enable OncoNano to clinically translate, the ultrasensitive pH probes and develop it from the lab bench through to establishing reproducible manufacturing of a stable formulation, to establishing safety and pharmacology, for an FDA IND filing. The specific aims in the SBIR Phase I, addresses the nanoprobe stability requirement, and addresses the nanoprobe sensitivity and specificity requirement in in-vivo models. The specific aims in Phase 2 of the project addresses the critical milestones on the path to clinical translation including- the scale-up requirement to meet the GLP and human testing needs, a demonstration of product safety in GLP in-vivo studies, culminating in an IND filing with the FDA.